Studies proposed in this application will evaluate the adjuvant activity of a natural peptide product when delivered using the needle-free method of intranasal mucosal immunization. We will build upon previous published studies from our group and utilize anthrax recombinant protective antigen (rPA) and a peptide immunogen to evaluate the natural peptide angiotensin-(1-7) (Ang-(1-7)) for its ability to provide nasal adjuvant activity for the induction of serum and mucosal antigen-specific humoral and cell- mediate immune responses to include serum IgG, mucosal IgA as well as systemic and mucosal epitope-specific CD8 responses. We will also initiate studies to better define the mechanism of action of this natural peptide adjuvant. Our specific aims are listed below and will utilize a mouse model of nasal immunization to perform early-stage preclinical studies to determine the safety and efficacy of Ang-(1-7) as a nasal vaccine adjuvant when compared to the gold-standard mucosal adjuvant cholera toxin (cholera toxin is not suitable for use in humans due to numerous toxicities). Information learned by the completion of the studies proposed in this application will provide a foundation for the development of a novel, natural peptide for use as an adjuvant for needle-free mucosal vaccines. Our specific aims are: Aim 1. Evaluate the nasal adjuvant activity of Ang-(1-7) when nasally delivered with protein or peptide immunogens. Studies performed in this aim will evaluate the adjuvant activity of Ang-(1-7) when nasally delivered with a protein antigen (anthrax protective antigen) or a peptide immunogen (C4-V3 HIV-1 peptide). When using the protein antigen, we will monitor vaccine-induced antigen-specific serum IgG, IgA and IgE responses as well as mucosal IgA. When using the peptide antigen, we will monitor the induction of epitope-specific CD8 responses using IFN3 ELISPOT and tetramer assays. The adjuvant activity of Ang-(1-7) will be compared to control adjuvants cholera toxin and CpG. Aim 2. Perform studies to evaluate the mechanism of action of Ang-(1-7) and its safety when used as a nasal vaccine adjuvant. Ang-(1-7) is known to signal via the MAS receptor. Studies performed in this aim will utilize the MAS antagonist A-779 to determine if the MAS receptor is required for the adjuvant activity of Ang-(1-7) and to determine if the use of Ang-(1-7) influences the cellular composition of the nasal cavity lymphoid tissue after nasal vaccination. Additional studies will evaluate the safety of Ang-(1-7) when used as a nasal vaccine adjuvant. PUBLIC HEALTH RELEVANCE: Studies proposed in this application will evaluate the angiotensin peptide Ang-(1-7) for its ability to be used as a vaccine adjuvant and enhance the immunogenicity of vaccines delivered by the nasal route. These vaccines are designed to be delivered intranasally and may someday replace vaccines injected with a needle. The identification and development of safe and effective nasal vaccine adjuvants is needed to make progress in the development of nasally-administered vaccines.